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Selective control of lignin biosynthesis in transgenic plants

Inactive Publication Date: 2002-09-12
RGT UNIV OF CALIFORNIA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0014] The invention additionally provides methods of enhancing lignification in a vascular plant by suppressing AGL8-like gene product expression in the vascular plant, whereby lignification is enhanced.

Problems solved by technology

Although lignins are essential for competitive survival of vascular plants, their resistance to degradation has had a negative impact on certain agricultural and industrial uses of plants.
Lignins are believed to limit forage digestibility by interfering with microbial degradation of fiber polysaccharides.
High lignin content also is problematic in the wood products industries, which contribute about 4% of the US Gross National Product and are an important component of the global economy.
In wood-pulp and paper industries, lignins are undesirable components that must be removed by costly chemical pulping.
Thus, both the digestibility of forage crops and the pulping properties of trees are adversely effected by high lignin content.
Yet, previous attempts to generate transgenic plants with altered lignin content have targeted biosynthetic enzymes and resulted in undesirable pleiotropic effects.

Method used

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  • Selective control of lignin biosynthesis in transgenic plants
  • Selective control of lignin biosynthesis in transgenic plants
  • Selective control of lignin biosynthesis in transgenic plants

Examples

Experimental program
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Effect test

example i

Production of a 35S::AGL8 Transgenic Arabidopsis Plant Displaying Reduced Lignification

[0134] This example describes methods for producing a transgenic Arabidopsis plant characterized by reduced lignification due to constitutive AGL8 expression.

[0135] Full-length AGL8 was prepared by polymerase chain reaction amplification using primer AGL8 5-.gamma. (SEQ ID NO:9; 5'-CCGTCGACGATGGGAAGAGGTAGGGTT-3') and primer OAM14 (SEQ ID NO:10; 5'-AATCATTACCAAGATATGAA-3'), and subsequently cloned into the SalI and BamHI sites of expression vector pBIN-JIT, which was modified from pBIN19 to include the tandem CaMV 35S promoter, a polycloning site and the CaMV polyA signal. Arabidopsis was transformed using the in planta method of Agrobacterium-mediated transformation essentially as described in Bechtold et al., C.R. Acad. Sci. Paris 316:1194-1199 (1993), which is incorporated herein by reference. Kanamycin-resistant lines were analyzed for the presence of the 35S-AGL8 construct by PCR using a prime...

example ii

Production of an Arabidopsis agl1 agl5 Double Mutant Displaying Reduced Lignification

[0140] This example describes the production of an agl1 agl5 double mutant displaying reduced lignification.

[0141] A. Production of an agl5 Mutant by Homologous Recombination

[0142] A PCR-based assay of transgenic plants was used to identify targeted insertions into AGL5 as described in Kempin et al., Nature 389:802-803 (1997), which is incorporated herein by reference. The targeting construct consisted of a kanamycin-resistance cassette that was inserted between approximately 3 kb and 2 kb segments representing the 5' and 3' regions of the AGL5 gene, respectively. A successfully targeted insertion produces a 1.6 kb deletion within the AGL5 gene such that the targeted allele encodes only the first 42 of 246 amino acid residues, and only 26 of the 56 amino acids comprising the DNA-binding MADS-domain. The recombination event also results in the insertion of the 2.5 kb kanamycin-resistance cassette wit...

example iii

Production of an agl8 Mutant Arabidopsis Plant Displaying Enhanced Lignification

[0157] This example describes methods for producing a non-naturally occurring plant characterized by enhanced lignification due to suppression of AGL8 expression.

[0158] A. Production of an agl8 Mutant

[0159] A mutation designated ful-1 was identified using large scale insertional mutagenesis with enhancer and gene trap Ds transposable elements (Sundarsen et al., supra, 1995; Springer et al., Science 268:877-880 (1995), each of which is incorporated herein by reference). This system utilized the maize Ac / Ds transposable elements and the reporter gene GUS. Transposition events were selected and screened for reporter gene expression patterns and mutant phenotypes. The ful-1 mutant was identified in the F3 progeny of an enhancer trap line. Backcrossing to wild type Landsberg erecta confirmed that ful-1 is a recessive mutation.

[0160] To address whether the ful-1 mutation was caused by insertion of the Ds-GUS e...

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Abstract

The present invention provides methods of selectively controlling lignin biosynthesis in plants such that lignification is reduced or enhanced, as desired. The invention provides, for example, a method of reducing lignification in a vascular plant by ectopically expressing a nucleic acid molecule encoding an AGL8-like gene product in the plant, whereby lignification is reduced due to ectopic expression of the nucleic acid molecule. An AGL8-like gene product useful in the invention can have, for example, substantially the amino acid sequence of an AGL8 ortholog such as Arabidopsis AGL8 (SEQ ID NO:2).

Description

[0001] This application is based on, and claims the benefit of U.S. Provisional Application No. 60 / 090,649, filed Jun. 25, 1998, which is herein incorporated by reference.BACKGROUND OF THE INVENTIONField of the Invention[0002] The present invention relates generally to agriculture and plant genetic engineering and more specifically to the production of genetically modified vascular plants in which the natural process of lignification is reduced or enhanced.Background Information[0003] Plant cell wall lignins (from the Latin lignum: wood) occur exclusively in higher plants and represent the second most abundant organic compound on the earth's surface after cellulose, accounting for about 25% of plant biomass. Cell wall lignification involves the deposition of phenolic polymers (lignins) on the extracellular polysaccharide matrix. The polymers arise from the oxidative coupling of three cinnamyl alcohols. The main function of lignins is to strengthen the plant vascular body, and the ri...

Claims

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Application Information

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IPC IPC(8): C12N15/82
CPCC12N15/8222C12N15/8223C12N15/8255
Inventor YANOFSKY, MARTIN F.LILJEGREN, SARAH
Owner RGT UNIV OF CALIFORNIA
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